Introduction to Stem Cell Principles and Biology
A stem cell is defined as an unspecialized cell that can both self-renew and give rise to differentiated progeny. In particular stem cells can divide to generate at least one cell that retains the stem cell identity, and can also give rise to progenitors, or precursor cells, which typically differentiate into tissue-specific cell types. Stem cells are derived from embryonic, fetal, or adult tissue and are broadly categorized accordingly. Recent advances in regenerative medicine support the development of new and emerging areas of integrative research including stem cells, gene- and cell-based therapies, and tissue engineering. The type of human cells, the use of growth factors and cytokines to stimulate the production, and growth and function of cells, along with the cell sources, have shown a significant therapeutic impact to date and represent a rapidly grown area of regenerative medicine.
KeywordsStem cells Embryonic stem cells Fetal stem cells Adult stem cells Regenerative medicine MSCs HSCs
Embryonic Stem Cells (ESCs)
Embryonic stem cells (ESCs) are derived from the inner cell mass of a blastocyst, which is formed 4–5 days after fertilization and exhibit the potential to self-renew without limit in culture. In more detail ESCs exhibit a high proliferation potential in vitro; maintain high levels of Oct-4 expression, telomerase activity, and a normal karyotype; and retain the potential to differentiate into cell types of all three lineages [1, 2].
Established human ESC lines were typically derived from in vitro fertilized embryos destined for destruction at in vitro fertilization units. In order to generate a single ESC line, 30–34 cells of the inner cell mass of blastocyst are isolated and expanded in vitro. Human ESC lines are cultured in growth medium supplemented with animal sera and maintained usually on mouse feeder layers (i.e., mouse embryonic fibroblasts) . Furthermore, ESCs are pluripotent with a great differentiation potential to various cell types. The differentiation potential of human ESCs can be evaluated either in vivo or in vitro, whereas ESCs can be cultured in vitro under certain culture conditions to induce differentiation into the desired cell type [1, 4, 5]. The in vivo models involve injecting cells into immunocompromised mice and analyzing the teratoma formation. However, it is notable that established ES lines may display some genomic instability . Thus, the use of ESCs for regenerative medicine is questioned, as ESCs appear to be tumorigenic and form teratomas that contain cell types representing all three primary germ layers in vivo . It is evident that, prior clinical use, it will be important to exclude undifferentiated stem cells from cell types or products derived from ESCs. Another important issue that remains unsolved and must be addressed is the immune rejection that these cells may provoke. It has been generally assumed that due to the fact that human ESCs and their differentiated derivatives can express high levels of major histocompatibility complex (MHC) class I antigens, any ES cell-based product will be subjected to graft rejection .
Induced Pluripotent Stem Cells (iPSCs)
In 2006, Yamanaka et al. managed to reprogram mouse skin fibroblasts into stem cells, similar to embryonic stem cells, by overexpressing four transcription factors OCT4, SOX2, KLF4, and c-MYC; these cells were characterized as induced pluripotent stem cells (iPSCs) . By using this approach, adult cells can be genetically reprogrammed to an embryonic stage by switching the expression of the necessary genes for the embryonic stem cell properties . The iPSCs have the ability to further differentiate into various types of cells, like ESCs, without the presence of concomitant ethical problems associated with the destruction of the blastocyst. Accordingly, Yamanaka and et al. managed to reprogram the “biological clock of the cell.” Since then, iPSCs have been generated from human somatic cells by using a variety of protocols. In subsequent studies, researchers replaced the original transcription factors with other combinations, but always in the presence of OCT-4, which represents an essential transcription factor for reprogramming somatic cells. The iPSCs resemble but are not identical to ESCs, as detailed genomic analysis reported the existence of epigenetic memory in iPSCs [7, 8].
To this end, iPSCs have been shown to possess some specific features or properties that can be acquired during the reprogramming process or are remnants of epigenetic modifications of the DNA derived from the parental tissue or cell that influence gene expression [7, 8]. These residual signatures of epigenomes and transcriptomes of the somatic tissue or cell of origin were termed as “epigenetic memory.” It has been reported that residual DNA methylation signatures derived from the somatic tissue of origin may favor their differentiation potential into lineages related to the donor cell while restricting alternative cell fates .
Advantages: (i) iPSCs are undifferentiated with unlimited differentiation potential into all cell types. In addition, iPSCs can be expanded in vitro to a high passage. These properties are allowing them to be used as a potential therapeutic tool in all tissues and organs . (ii) Studying iPSCs derived from pathological or normal tissue can offer a better understanding of a disease and the relevant molecular pathways. iPSCs are often termed as a “disease in a dish” . (iii) No ethical considerations are related to iPS generation.
Disadvantages: (i) The efficiency or reprogramming is very often low and depends on the donor tissue and the reprogramming method. (ii) Prior transplantation into patients, it is needed to ensure that iPSCs are fully differentiated into the required specialized cells. (iii) iPSCs, like ESCs, are reported to form teratomas in vivo after transplantation . (iv) Epigenetic memory in iPSCs influences the gene expression .
Fetal Stem Cells (FSCs)
Fetal stem cells represent a relative new source of stem cells. These cells can be derived either from the fetus or from the supportive extraembryonic structures. FSCs have been recently isolated from several tissues such as amniotic fluid, amnion, umbilical cord blood, Wharton’s jelly, placenta, fetal liver or fetal bone marrow [11, 12, 13, 14].
Recent reports describe fetal stem cells as ideal cell types for regenerative medicine because they (i) are easily accessible as these cells are usually derived from tissues that are normally discarded following birth, such as umbilical cord, placenta, or amnion, (ii) exhibit high proliferation rates in vitro, (iii) do not form teratomas when injected to immunosuppressed mice in vivo, (iv) do not present ethical reservations like embryonic stem cells (ESCs) and (iiv) exhibit functional features indicating that they represent intermediates between ESCs and adult stem cells (e.g., amniotic fluid stem cells express the pluripotency marker Oct-4 in high levels [15, 16]). Another important issue is that early fetal stem cells appear to have pre-immune status and can be used with limited implications compared to adult stem cells in allogenic transplantations. In particular, these cells do not express HLA-class II, but express HLA-class I antigens, and they do not elicit lymphocyte proliferation in vitro [11, 12, 13].
However, these cells have a limited differentiation potential compared with ES cells, as they cannot give rise to all cell types of the three germ layers. It will remain necessary to show that fetal stem cells can differentiate into fully functional committed cells in vivo in order to evaluate better their therapeutic potential [11, 14].
In the following sections, fetal sources such as amniotic fluid, umbilical cord blood and extraembryonic tissues will be analyzed in detail.
Amniotic Fluid (AF)
AF serves as a protective liquid for the developing embryo, providing mechanical support and the required nutrients during embryogenesis. Amniocentesis has been used for many decades as a routine procedure for fetal karyotyping and prenatal diagnosis, allowing the detection of a variety of genetic diseases.
AF also represents a rich source of a stem cell population deriving either from the fetus or the surrounding amniotic membrane. Additional investigations by several groups have been recently focused on the cellular properties of amniotic-derived cells and their potential use in preclinical models and in transplantation therapies [12, 16, 17, 18, 19].
The amniotic fluid cells (AFCs) represent a heterogeneous population derived from the three germ layers. These cells share an epithelial origin and are derived from either the developing embryo or from the inner surface of the amniotic membrane, which are characterized as amniotic membrane stem cells. The AFCs are mainly composed of three groups of adherent cells, categorized based on their morphological, growth, and biochemical characteristics. Epithelioid (E-type) cells are cuboidal to columnar cells derived from the fetal skin and urine, amniotic fluid (AF-type) cells are originating from fetal membranes, and fibroblastic (F-type) cells are generated mainly from fibrous connective tissue. Both AF- and F-type cells share a fibroblastoid morphology, and the dominant cell type appears to be the AF-type, co-expressing keratins and vimentins. Several studies have documented that human amniotic fluid stem cells (AFSCs) can be easily obtained from a small amount of second trimester AF, collected during routine amniocenteses, a procedure with spontaneous abortion rate ranging from 0.06% to 0.5%. Up to date, a number of different cultivation protocols have been described, leading to enriched stem cell populations [12, 16, 20, 21]. The isolation of AFSCs and the respective culture protocols were summarized in a review by Klemmt et al.  and can be categorized as follows: (i) a single-step cultivation protocol, where the primary culture was left undisturbed for 7 days or more until the first colonies appear; (ii) a two-step cultivation protocol, where amniocytes, not attached after 5 days in culture, were collected and further expanded; (iii) cell surface marker selection for CD117(c-kit receptor); (iv) mechanical isolation of the initial mesenchymal progenitor cell colonies formed in the initial cultures; and (v) short-term cultures to isolate fibroblastoid colonies. The majority of the AFSCs, isolated following these methodologies, shared a multipotent mesenchymal phenotype and exhibited higher proliferation potential and a wider differentiation potential compared to adult MSCs . The AFSCs exhibit a typical mesenchymal marker expression profile, being positive for markers such as CD90, CD73, CD105, CD29, CD166, CD49e, CD58, and CD44, as determined by flow cytometry analyses. Additionally, these cells expressed the HLA-ABC antigens, whereas the expression of the hematopoietic markers CD34 and CD45, the endothelial marker CD31, and the HLA-DR antigen was undetected. More importantly, the majority of cultured AFSCs expressed pluripotency markers, such as the octamer-binding protein 3/4 (Oct-3/4), the homeobox transcription factor Nanog (Nanog), and the stage-specific embryonic antigen 4 (SSEA-4) [16, 18, 19].
It was also reported that amniocyte cultures contain a small population of CD117 (a tyrosine kinase specific for stem cell factor present primarily in ESCs and primordial germ cells)-positive cells that can be clonally expanded in culture. The differentiation properties of CD117+ AFSCs were tested for the first time in vivo, proving in this way their stem cell identity. Experimental evidence suggested that AFSCs are derived from spindle-shaped fibroblastoid cells .
In an attempt to analyze the AFSCs subpopulations, our group recently identified two morphologically distinct populations of AFSCs of mesenchymal origin, with different proliferation and differentiation properties, termed as spindle shaped (SS) and round shaped (RS). Both subpopulations were expressing mesenchymal stem cell markers at similar levels. However, it was identified that SS colonies expressed higher levels of CD90 and CD44 antigens compared to RS colonies .
Umbilical Cord Blood
Umbilical cord blood was first seen as biological waste product post birth. In 1980, Di Landro et al. reported the colony-forming capacity of cord blood to be similar to bone marrow . In 1988, the first successful cord blood transplant took place in France for Fanconi’s anemia with the donor being an identical human leukocyte antigen (HLA)-matched sibling by Eliane Gluckman . Most importantly, the patient is reported to be alive and well 18 years after the transplantation .
In 1990, three more patients had been transplanted for Fanconi’s anemia, and it was reported that cord blood transplantation may also be applicable to other diseases with a possibility of transplanting adults. The cord blood cellular product represented an advantageous source of hematopoietic stem/progenitors cells for transplantation.
In 1991, the first report of a Public Cord Blood Bank for unrelated cord blood transplants and in 1992 was published, reporting the characterization of cord blood by flow cytometry by Dr. Gluckman’s team. This study demonstrated that the cord blood graft cells represented both suppressive and naive cells .
Cord blood cells are considered a gold standard product for hematopoietic transplantation and reconstitution and a potential product for regenerative medicine . In addition, hematopoietic cell transplantation is a gold standard worldwide for a long list of hematopoietic diseases, which includes leukemia; myelodysplastic syndrome; myeloproliferative and lymphoproliferative disorders; inherited metabolic, immune, or platelet disorders; and other malignancies. Transplantation of umbilical cord blood is characterized by low immunogenicity as indicated by reduced acute GVHD [14, 27, 28, 29].
Cord blood in the past was considered as a product for transplantation mainly in children due to the low number of cells that could be harvested from a single cord blood collection. However, recently adult cord blood transplantation has been successfully studied using double cord blood unit transplantation. Further it has been demonstrated that UCB can provide long-term hematopoietic reconstitution in adults [26, 27, 28].
Stem Cells Derived from Extraembryonic Tissues
Amniotic Membrane (AM)
The amniotic membrane , lacking any vascular tissue, forms most of the inner layer of the fetal membrane and is composed of (i) an epithelial monolayer consisting of epithelial cells, (ii) an acellular intermediate basement layer, and (iii) an outer mesenchymal cell layer, rich in mesenchymal stem cells and placed in close proximity to the chorion . AM was used in clinic for many decades for wound healing in burns, promoting epithelium formation and protecting against infection. Recently, the use of AM has been evaluated as a wound dressing material for surgical defects of the oral mucosa, ocular surface reconstruction, corneal perforations, and bladder augmentation. Amniotic membrane stem cells (AMSCs) include two types, the amniotic epithelial cells (AECs) and the amniotic membrane mesenchymal stromal cells (AM-MSCs) derived from the amniotic epithelial and the amniotic mesenchymal layers, respectively [13, 14]. Both cell types are originated during the pre-gastrulation stages of the developing embryo, before the delineation of the three primary germ layers, and are mostly of epithelial nature. A variety of protocols has been established for AECs and AM-MSCs isolation, primarily based on the mechanical separation of the AM from the chorionic membrane and the subsequent enzymatic digestion. AM-MSCs exhibited plastic adherence and fibroblastoid morphology, while AECs displayed a cobblestone epithelial phenotype. AM-MSCs shared similar phenotypic characteristics with the ones derived from adult sources. More interestingly, AM-MSCs exhibited a higher proliferation rate compared to MSCs derived from adult sources and a multilineage differentiation potential into cells derived from the three germ layers [13, 14, 30].
The placenta serves the functions of all organs of the developing embryo by working in association with the mother. During pregnancy, it functions as the embryo’s lungs, kidneys, digestive system, liver, and immune system. It is evident that due to the placenta, an embryo can survive until birth, even when one or more vital organs fail to develop.
The placenta also serves to protect the developing embryo from an attack by the mother’s immune system, since the embryo and the placenta are genetically unique and distinctly different from the mother [13, 14].
Several stem cell populations are derived from the placenta with the best studied the placenta mesenchymal stem/stromal cells (MSCs). Placenta MSCs express markers of pluripotency such as SEEA-4, Oct-4, Stro-1, and Tra 1-81, and also they have a wide range of differentiation potential. It has been reported that placenta MSCs are capable of in vivo differentiation into neuronal, glial, and insulin-positive cells and hepatocytes and the generation of heart valves seeded in scaffolds [13, 14, 17, 20, 21].
Adult Stem Cells
Adult stem cells are found, in small percentage, in almost all tissues after birth and are able to self-renew and differentiate, in most cases, into cell types of the tissues that they originate . However, recent studies have identified adult stem cells with a greater range of potential than that originally believed. The most well-characterized adult stem cell types are the ones derived from human bone marrow (BM). However, adult stem cells have been also isolated from the blood, brain, fat, liver, muscle or pancreas .
Since adult stem cells are often a very small percentage of the total cells of a tissue or organ, isolation and expansion are considered difficult and time-consuming. In many cases, investigators isolate adult stem cells based on their surface antigen expression or by examining their differentiation potential. In some cases, the lack of a single marker for their characterization leads to the isolation of a heterogeneous population with questioned stem cell identity .
As standardized protocols develop for adult stem cell isolation, more rigorous criteria will develop for determining true stem cell populations and their differentiation potential.
ΒΜ stem cells represent the most well-characterized example of adult stem cells, are fairly easy to isolate, and have been the most thoroughly investigated, with several reports demonstrating their contribution to regenerative medicine. ΒΜ stem cells consist of hematopoietic stem cells (HSCs), which can give rise to blood cell lineages and endothelial progenitor cells (EPCs), and mesenchymal stromal cells (MSCs), which have been shown to differentiate into mesodermal phenotypes (adipocytes, chondrocytes, and osteocytes). HSCs and MSCs can also be derived in high numbers from umbilical cord blood and Wharton’s jelly, respectively. Although adult stem cells may represent a valuable tool for autologous transplantations, their proven multilineage differentiation potential is limited . As examples of adult stem cells, HSCs, MSCs and neural stem cells are described in more detail in the following sections.
Hematopoietic Stem Cells (HSCs)
Hematopoietic stem cells (HSCs) represent the stem cell population responsible for the development, maintenance and regeneration of the blood-forming tissue during life. Because HSCs can reconstitute and restore the hematopoietic system of a myeloablated host, they have been used for treating hematologic disorders, starting in 1945 .
HSC presence has been shown in adult mouse bone marrow as a cell population marked by c-kitpos, thy-1low and sca-1pos . Human HSCs are characterized by c-kitpos, Thy-1pos and CD34pos expression. HSCs from mice and humans are being isolated, starting with a lineage depletion step in which lineage-specific cells (B220, CD3, 4, 8, 11b Mac-1, Gr-1 and Tcr-119 for mice and CD10, 14, 15, 16, 19, and 20 in human) are depleted. The resultant population, termed as Linneg, can be enriched and is able to repopulate the bone marrow of a lethally irradiated host [33, 34].
HSCs can be in vitro expanded by co-culturing them with bone marrow-derived stromal cells. Several subpopulations within Linneg HSCs are existing. One such homogenous population is characterized as side population (SP) cells based on their unique ability to exclude Hoechst dye. This subpopulation can be examined by FACS analysis and SP cells fall within a separate population to the side of the rest of the cells on a dot plot of emission data. These cells are also able to home and engraft to the BM of a lethally irradiated host .
HSCs, which primarily reside in the BM, maintain blood formation and replenish themselves throughout the adults’ lifespan. The activity of bone marrow HSCs was discovered in 1960s, identifying a robust contribution of donor BM cells in lethally irradiated recipient mice. After 30 years of work, the contribution of donor hematopoietic cells in recipients had been demonstrated to derive from a few specific “clones,” suggesting the existence of HSCs . HSCs was isolated for the first time in 1988 when Weissman et al. enriched HSCs from the murine BM using a fluorescent-activated cell sorter . Since then, numerous groups have demonstrated that HSCs possess stem cell properties including the ability to self-renew as well as to differentiate into all of the hematopoietic lineages [33, 34]. HSCs are committed to a differentiation program, in that they exclusively create all of the cells of the hematopoietic origin. Clinical trials of HSC transplantation for the purpose of restoring hematopoiesis have been widely performed for treating leukemia, severe autoimmune disease, and severe combined immunodeficiency. In addition, HSC treatment has been used in adjunct to chemotherapy for other cancers such as breast cancer, neuroblastoma and testicular cancer. Currently, HSC research is studying the development of novel ways to improve the transplantation success by reducing graft-versus-host disease and infection during recovery and accelerating hematopoiesis after transplantation [14, 33, 34].
Mesenchymal Stromal Cells (MSCs)
The first descriptions of fibroblastic cells that could be isolated and grown from bone marrow and also retained the ability to differentiate to bone tissue were presented by Alexander Friedenstein in the 1960s, using guinea pig bone marrow as the source. Friedenstein used the term “osteogenic cell” in order to describe the properties of these cells. MSCs can differentiate into osteocytes, adipocytes and chondrocytes, but they can also exhibit multilineage in vitro differentiation depending on their source of origin (fetal sources of MSCs have been characterized during the last 20 years and are described earlier in this chapter) .
Human MSCs were firstly isolated from BM by their adherence to tissue culture plastic vessel and were expanded through multiple passages in medium containing high concentrations of fetal calf serum (FCS). However, the proliferation rates and other properties of the cells gradually change during expansion. The cells are cloned as single-cell-derived colonies, but both the colonies and the cells within a colony, are heterogeneous in morphology, rates of proliferation and efficacy with which they differentiate [36, 37, 38].
Adherence to plastic in standard culture conditions (expandability of these cells without losing their differentiation potential)
Phenotype: Positive for CD105, CD90, and CD73 and negative for CD45, CD34, CD14 or CD11b, CD79a or CD19, and HLA-DR
In vitro differentiation: osteoblasts, adipocytes, and chondroblasts (demonstrated by staining of in vitro cell culture)
The current ISCT definition recommends to use the term “multipotent mesenchymal stromal cells” (MSC) instead of “mesenchymal stem cells.” However, literature review showed that after 2006, ISCT members (including the authors of MSC position paper) themselves frequently use terms “mesenchymal stromal cells” or “mesenchymal stem cells.”
MSCs produce a number of secreted factors such as vascular endothelial growth factor (VEGF), stem cell factor (SCF-1) , leukemia inhibitory factor (LIF), granulocyte colony-stimulating factor (G-CSF) , macrophage colony-stimulating factor (M-CSF), granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukins (IL-1, IL-6, IL -7, IL -8, IL -11, IL -14, and IL -15), stromal cell-derived factor (SDF-1), Flt-3 ligand, and others. The expression of these factors may be modulated through interactions with other cell types . Due to the secretion of the various types of factors, Caplan in 2011 used the term “drug store” to describe these cells .
MSCs also exhibit homing properties to sites of tissue injury, particularly ischemic regions of the heart where the MSCs may prevent deleterious remodeling. In addition, MSCs also have the ability to alter immune responses and engraft in allogeneic recipients, and it has been reported that the MSC treatment has been used to clinically treat graft-versus-host disease (GVHD) [14, 40].
Neural Stem Cells (NSCs)
Neural stem cells (NSCs) represent the most primitive and uncommitted cells of the nervous system. Evidence support that these cells give rise to the vast majority of more specialized cells of the central nervous system (CNS) and peripheral nervous system (PNS). The term “neural stem cell,” is in contrast to the term “progenitor” cell (i.e., describes cells that are lineage committed to give rise to only one category of neural cell type, such as glial cells, neurons, etc.) .
Neural stem cells must be capable of (i) generating all neural lineages (neurons, astrocytes and oligodendrocytes), (ii) having limited capacity for self-renewal and (iii) being able to give rise to cell types in addition to themselves through asymmetric cell division [14, 40].
Neural Stem Cells in the Developing Brain
During embryogenesis, CNS development initiates with the induction of the neuroectoderm, which forms the neural plate and then the neural tube. Within these neural structures, a complex and heterogeneous population of neuroepithelial progenitor cells (NEPs) represents the earliest neural stem cell type. As CNS development proceeds, NEPs generate distinct neural stem/progenitor populations. NEPs also undergo symmetric divisions to expand NSCs. In the later stage of neural development, it has been reported that NSCs undergo asymmetric divisions and differentiate into lineage-restricted progenitors. Thus, intermediate neuronal progenitor cells are formed that subsequently differentiate into neurons. Previously it was stated that neurogenesis in the adult mammalian CNS was complete, implying incapability of mitotic divisions in order to generate new neurons and therefore lacking in the ability to restore or regenerate damaged tissue caused by diseases (such as Parkinson’s disease, multiple sclerosis) or injuries (such as spinal cord and brain ischemic injuries). However, recent strong in vivo and in vitro evidence support that NSCs exist in the mature mammalian CNS [14, 42].
Mason and Dunnill in their review in 2008  summarized the clear distinction between organ regeneration and organ repair. Regenerative medicine includes activities such as surgery, surgical implants, and biomaterial scaffolds. Thus, regenerative medicine integrates human cell therapy, gene-based methods, biomaterials and molecular medicine. To this end, regeneration is described as “the process in humans, whereby lost specialized tissue is replaced by proliferation of undamaged specialized cells”, whereas repair is “the replacement of lost tissue by granulation tissue, which matures to form scar tissue” . Consequently, repair is an adaptation to loss of normal organ mass leading to restoration. It is evident that regeneration restores the normal structure and function of the organ, whereas repair does not. Therefore, regenerative medicine replaces or regenerates human cells, tissues or organs to restore or establish normal function .
The major aim of regenerative medicine is to establish novel therapies for severe injuries or chronic diseases in which patients’ own responses do not suffice to restore functional tissue. Recent reports describe several major medical needs, which might be addressed by regenerative technologies. Such examples include severe burns, spinal cord injuries, congestive heart failure, diabetes, Alzheimer’s and Parkinson’s diseases and others .
The areas of specialty of regenerative medicine continue to change rapidly; however, the main focus of regenerative medicine therapies is the use of stem/progenitor cells into clinical applications for both allogenic and autologous transplantations. The field now integrates a wide area of scientific fields and technologies such as stem cells, genetic reprogramming, gene therapy, nuclear transfer, genomics, proteomics, cloning and tissue engineering. The extension of novel therapeutic areas, including organ generation with 3D structure, depends on scaffold engineering, material science and/or bioreactor technology [14, 43].
Cells for structural repair or replacement (e.g., cultured dermal fibroblasts as skin replacement or chondrocytes for repair of cartilage)
Cells for correction of a physiological or metabolic problem
Autologous cells are derived from the patient to be treated, whereas allogeneic cells are derived from a donor. Several recent studies described allogeneic cell therapies developed including cultured keratinocytes as dermal matrices for the repair of cutaneous wounds, hepatocytes for liver repair, pancreatic islets for diabetes and hematopoietic stem cells for bone marrow transplantation in leukemia and other types of cancers [14, 43, 44].
Understanding the nature of the problem that need treating, the role that the cell can play in solving the problem (i.e., engraftment and differentiation at the site of the injury or paracrine effect) and also the related molecular mechanisms are critical to develop a successful cell therapy . Scientists in regenerative medicine have strived to understand the interaction of cells, extracellular matrices and biological factors in order to develop tissue-engineered products for repair and replacement of injured tissues.
However, there are several limitations related to the type of cells that can be isolated, the condition of the tissue, the patient’s age, and others. To achieve this goal, extended in vitro assays and in vivo animal models are needed to recapitulate the molecular events that take place and understand the mechanisms underlying the interactions of cells, extracellular matrices, and biological factors  (Fig. 1).
To date, regenerative medicine introduces novel methods and strategies to replace or regenerate cells, tissue or organs in order to restore and establish normal function. These strategies mostly include cell-based therapies combined with the use of biomaterials and scaffolds. The characterization and the basic properties of stem/progenitor cell types are crucial in respect of their use in potential clinical applications. Most importantly, the type and the source of cells remain of central focus for the approaches adopted in cell-based therapies. A primary issue remains the choice between using patient’s own cells or cells derived from allogenic donors.
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